The CDNA Cloning Of Bovine Neutrophil Cell β-Defensins And Construction Of Its Prokaryotic Expression Plasmids

A specific pair of primers were designed and synthesized according to the sequence of bovineβ-defensin gene published in GenBank. Total cellular RNA was purified from the Bovine Neutrophil cell with the total RNA isolation kit. Then the Bovineβ-defensin cDNA fragment was amplified by reverse transcription-polymerase chain (RT-PCR), the RT-PCR products were inserted into the pMD18-T cloning vector. Then the cDNA fragment of interest was sequenced. The result showed that the obtained 189 bp DNA fragment is identical with the BNBD-7 sequence registered in GenBank. The cDNA fragment code 63 amino acid residues.Compared with the gene published in GenBank,the cDNA identity was 96.3%.The products were inserted into the vector pET-32a and pQE-30 to construct prokaryotic Expression Plasmids pET-32a/ BNBD-7 and pQE-30/ BNBD-7 have laid a foundation for later expression,biological function and activity study ofβ-defensin.